Zinc-dependent modulation of ceftriaxone killing in tolerant Neisseria gonorrhoeae

Ceftriaxone tolerance in Neisseria gonorrhoeae is increasingly recognized as a phenotype that allows prolonged bacterial survival at inhibitory drug concentrations without altering minimum inhibitory concentrations (MICs). Host environmental factors are likely to influence such tolerance phenotypes, but experimental evidence remains limited. Zinc is a host-relevant transition metal that shapes bacterial stress responses and ribosomal function. Transcriptomic analyses have previously linked ceftriaxone tolerance in N. gonorrhoeae to a zinc-sparing ribosomal programme. In this study, we examined whether zinc availability modifies ceftriaxone killing kinetics in a paired tolerant and non-tolerant clinical isolate of N. gonorrhoeae. Both isolates had identical ceftriaxone MICs of 0.008 μg/mL. Using time-kill assays, we show that zinc supplementation attenuates early ceftriaxone-mediated killing in both phenotypes, resulting in shallower killing slopes and increased areas under the killing curve between 2 and 8 h after drug exposure. Zinc was associated with transiently higher viable counts during the mid-phase of killing, with a numerically greater effect observed in the tolerant isolate. Despite these differences in early and mid-phase dynamics, all ceftriaxone-exposed cultures were at or below the limit of detection by 24 h, irrespective of zinc availability. Within the limitations of this in vitro study using a single paired tolerant and non-tolerant isolate, these findings provide functional evidence that zinc availability can act as a conditional modifier of ceftriaxone killing dynamics in N. gonorrhoeae, influencing short-term survival without altering MICs or final outcomes.